Flow cytometry represents one of the essential tools employed in such fields as cytologic biology, cellular immunochemistry, and cytodiagnosis for cancer detection. Essentially, it aims at the classification of cells according to their sizes, types, contents of intracellular components, and like characteristics. Flow cytometry involves the pigmentation of cells with fluorescent dyes. The pigmented cells are caused to individually fluorescence under laser beam irradiation while flowing through slender tubing. The intensities of fluorescence of the individual cells are measured for the determination of their sizes, relative amounts of DNA (deoxyribonucleic acid), etc. Flow cytometry analysis can identify and enumerate lymphocyte subsets in human cells in suspension.
One usage of the system, apparatuses and devices according to the present invention can be found in immunology and pathology laboratories at hospitals all over the world.
Conventionally, such cell pretreatment has been mostly performed manually at the cost of much time and labor. Manual pretreatment is also undesirable by reasons of unavoidable human errors and the non-uniformity of operations from one individual operator to another. The advent of an apparatus capable of full automation of cell pretreatment has thus been long awaited by the cytologists for the elimination of human toil and for gaining stability and constancy in the operations involved. No such system or apparatus is known, because of the complexities of the processes of cell pretreatment. However, there are several companies trying to partially automate some of the processes.
Becton-Dickinson (BD) with its BD Facs Lyse Wash Assistant (for automatic lysing, washing and centrifuging) and Beckmann Coulter with its TQ Prep Workstation (for automatic lysing procedure), the PrepPlus 2 Workstation (for pipetting of reagents, patient samples, controls and fluorospheres into daughter tubes), and the Antibody Cocktail Preparation Workstation (for cocktail mixing) offer complementary automation to the manual process.
The BD Facs Lyse Wash Assistant does not pipette reagents or patient samples. The centrifuge solution is based on a single vial (tube) centrifuge, where the vial rotates around its own axe. The inconvenience with that centrifuge solution is that the cells in the sample won't gather in the bottom of the vial, but on the walls. One lab engineer interviewed mentioned that there is a problem with losing too many cells in this process.
EP 0 645 631 B1, assigned to Becton Dickinson, describes an automated system for preparing samples and for loading the samples to an analyzer, wherein the system includes a sample carousel which mixes and indexes a plurality of sample tubes to a sample aspiration station.
Beckman Coulter offers several instruments in order to automate the complete manual procedure. The unfortunate outcome of using that strategy is that the automated process will still include several manual activities after buying these instruments. After pipetting patient samples and reagents in the PrepPlus 2 Workstation, the sample(s) must be manually moved to the TQ Prep Workstation for lysing procedure. The FP1000 Cell Preparation System is also a part of the Beckman Coulter family and is being used for lysing solutions.
EP 1 468 266 A1, assigned to Beckman Coulter, relates to an adjustably controllable environment containment system for a flow cytometer to protect both the particles being processed by the flow cytometer from contamination by both biological and non-biological materials and to protect persons using the flow cytometer from being exposed to the particles being processed and the chemistry utilized in the processing of such particles.
JP 2010133727, assigned to Beckman Coulter, regards a cleaning mechanism for cleaning an approximately cylindrical dispensation nozzle.
US 2011/017238 A1, assigned to Beckman Coulter, teaches a nozzle cleaning method and a nozzle cleaning device which allow to surely perform cleaning of a dispensing nozzle and which allow reduction in cleaning time. For this purpose, a nozzle cleaning method for cleaning a dispensing nozzle for suctioning and discharging a liquid includes: a first cleaning step in which, after termination of dispensing, an inner wall surface of the dispensing nozzle is cleaned in an upper portion of a storage tank overflowed with a cleaning liquid by discharging a liquid for preload; and a second cleaning step in which at least an outer wall surface is cleaned by lowering and immersing the dispensing nozzle into the storage tank overflowed with the cleaning liquid.
EP 2 407 791 A1, assigned to Beckman Coulter, relates to an analyzer comprising: a reading section for storing or obtaining specimen information including a sample type of a specimen, and specimen container information; a liquid level detecting section for detecting a liquid level and/or an interface of the specimen; a dispensing apparatus for dispensing a specimen; a washing apparatus for washing a dispensing probe; a calculating section for calculating a contamination adhering range of an external wall surface of the dispensing probe, based on the sample type, specimen suction-position and specimen container information stored or obtained by the reading section, as well as liquid level and/or interface information of the specimen detected by the liquid level detecting section; and a washing control section for controlling a washing range based on the contamination adhering range.
WO 2010/110265 A1, assigned to Beckman Coulter, describes dispenser comprising a storage section for storing a voltage correction coefficient for each specimen type and voltage correction coefficients based on the type of containers which contain specimens, an information reader for acquiring specimen information and container information, a calculating section for calculating a threshold voltage on the basis of the voltage correction coefficient of a specimen and the voltage correction coefficient of a container, which have been extracted from the storage section, and a determination section for determining to be liquid-level detection when the signal received by a dispensing probe outputs the threshold voltage for a predetermined period or longer.
U.S. Pat. No. 5,030,554, assigned to Beckman Coulter, discloses a method for rapid preparation of a whole blood sample for photo-optical analysis.
EP 0 418 026 A2 refers to an apparatus for pretreating cells for flow cytometry.
WO 2009/150632 A2 discloses an apparatus for preparing controlled amounts of liquid for cytometry, which apparatus comprises a sampler having: motion imparting means for moving a main tray with one or more tubes thereon, each containing a controlled amount of liquid; a support and moving unit for supporting and moving a syringe that transfers preparation liquids into the tubes; a motion imparting mechanism for moving the piston of the syringe. The apparatus further comprises a centrifuge located beside the sampler and having an access opening for receiving the tubes, for selective removal of residues to be discarded from the controlled amounts of liquid. Furthermore, the sampler comprises a motorized gripping mechanism disposed in the support and moving unit and coaxial with the syringe, the support and moving unit being movable to allow the tubes to be carried from the main tray to the centrifuge, and vice versa.
EP 0 628 822 A2 teaches a blood analysis system or instrument, generally, including an incubator station, a sample and reagent holding station, a pipette assembly, a centrifuge, an analysis station, and a transport assembly. Generally, the incubation station holds containers while reagents and fluids are being dispensed in those containers, and, if desired, for incubating the containers. The sample and reagent holding station holds samples and a plurality of reagents, and the pipette assembly transfers fluids from that sample and reagent holding station to containers in the incubation station. The centrifuge is provided for centrifuging the container, and the analysis station is provided to analyze the containers, optically to identify reactions therein. The transport assembly carries the containers between the incubator station, the centrifuge, and the analysis station. Preferably, the pipette assembly is automatically operated to draw fluids and preselected reagents from the sample and reagent holding station, and to dispense fluids into the containers held in the incubation station to produce predetermined solutions therein. Also, the transport subassembly is automatically operated to carry containers from the incubator station to the centrifuge after the predetermined solutions have been produced in the containers, and then to carry the containers from the centrifuge to the analysis station.
None of the previously mentioned prior art products teaches a fully automated system as well as its sufficiently developed components/units/devices therein, where the system and its different components/devices are being adapted for cooperating and working together in a streamlined and seamless manner and for preparing of cells that are to be thereafter used in flow cytometry analysis.
One of the problems left unsolved in the art of flow cytometry is how to expedite the complete process of pretreating cells to be studied. Such pretreatment consists of many steps to be followed strictly in a prescribed order. Among the steps are introduction of reagents into cell samples within sample tubes, centrifugal treatment of the sample-reagent mixtures, removal of the unnecessary liquid tops from the sample tubes, staining of the cells with a fluorescent dye, and filtration of the samples. The actual process is much more complex.
One aspect of the present invention is to provide a system and improved apparatuses/devices for a fully automated and streamlined, as well as more effective/efficient, preparation or pretreatment procedure of cells preparatory to their flow cytometric or similar cytological studies.
It is also an aspect of the present invention is to provide for reducing or eliminating any possibility for contamination of cell sample(s) being prepared or pretreated.
Another aspect of the present invention is to provide for reducing the loss of too many cells during the preparation or pretreatment procedure or process.
Yet another aspect of the present invention is to provide for full control of antibody and reagent usage, as well as minimizing faulty on analysis due to manual mistakes when e.g. staining.